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Background: Alkali production via arginine deiminase system (ADS) of oral bacteria plays a significant role in oral ecology, pH homeostasis and inhibition of dental caries. ADS activity in dental plaque varies greatly between individuals, which may profoundly affect their susceptibility to caries., Objective: To investigate the effect of arginine on the growth and biofilm formation of oral bacteria., Methods and Results: Polymicrobial dental biofilms derived from saliva were formed in a high-throughput active attachment biofilm model and l-arginine (Arg) was shown to reduce the colony forming units (CFU) counts of such biofilms grown for various periods or biofilms derived from saliva of subjects with different caries status. Arg hardly disturbed bacterial growth of Streptococcus mutans, Streptococcus sobrinus, Streptococcus sanguinis and Streptococcus gordonii in BHI medium, but only inhibited biofilm formation of S. mutans. Scanning electron microscope (SEM) showed S. mutans biofilms harboured fewer cells grown with Arg than that without Arg, even in the initial 2h and 8h phase. Confocal laser scanning microscope (CLSM) images of poly-microbial dental and S. mutans biofilms revealed the biofilms grown with Arg had lower exopolysaccharide (EPS)/bacteria ratios than those without Arg (P=0.004, 0.002, respectively). Arg could significantly reduce the production of water-insoluble EPS in S. mutans biofilms (P<0.001); however, quantitative real-time PCR (qRT-PCR) did not show significantly influence in gene expression of gtfB, gtfC or gtfD (P=0.32, 0.06, 0.44 respectively)., Conclusions: Arg could reduce the biomass of poly-microbial dental biofilms and S. mutans biofilms, which may be due to the impact of Arg on water-insoluble EPS. Considering the contribution to pH homeostasis in dental biofilms, Arg may serve as an important agent keeping oral biofilms healthy thus prevent dental caries., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Objectives: In vitro methods to study dental biofilms are useful in finding ways to support a healthy microbial balance in the oral cavity. The effects of sucrose, xylitol, and their combination on three strains of Streptococcus mutans and one strain of Streptococcus sobrinus were studied using a dental simulator., Methods: A simulator was used to mimic the oral cavity environment. It provided a continuous-flow system using artificial saliva (AS), constant temperature, mixing, and hydroxyapatite (HA) surface in which the influence of xylitol was studied. The quantities of planktonic and adhered bacteria were measured by real-time qPCR., Results: Compared against the untreated AS, adding 1% sucrose increased the bacterial colonization of HA (p<0.0001) whereas 2% xylitol decreased it (p<0.05), with the exception of clinical S. mutans isolate 117. The combination of xylitol and sucrose decreased the bacterial quantities within the AS and the colonization on the HA by clinical S. mutans isolate 2366 was reduced (p<0.05). Increasing the concentration (2%-5%) of xylitol caused a reduction in bacterial counts even in the presence of sucrose., Conclusions: The continuous-culture biofilm model showed that within a young biofilm, sucrose significantly promotes whereas xylitol reduces bacterial colonization and proliferation. The results indicate that xylitol affects the ability of certain S. mutans strains to adhere to the HA. Clinical studies have also shown that xylitol consumption decreases caries incidence and reduces the amount of plaque. This study contributes to the understanding of the mechanism behind these clinical observations., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Objective: The aim of this study was to determine the antibacterial effect of different formulations containing Lysozyme and Lactoferrin and drug delivery system as well as poloxamer 407 with the trade name of Pluronic F-127 and/or freeze dried liposome containing DOTAP [freeze dried Liposomal DOTAP] on Streptococcus sobrinus, Streptococcus mutans and Lactobacillus acidophilus in comparison with 0.2% chlorhexidine., Materials and Methods: The antibacterial effect was assessed by determining the minimum inhibitory concentrations (MIC) for the study and control groups on Streptococcus sobrinus, Streptococcus mutans and Lactobacillus acidophilus. The amounts of biofilm formation accumulation of Mutans Streptococci for 24h on sterile hydroxyapatite discs after application of different formulations were evaluated. The different formulations studied were: (1) Sorensen's Buffer Solution, (2) a gel formulation containing only poloxamer 407, (3) Lysozyme and Lactoferrin dissolved in Sorensen's Buffer Solution, (4) poloxamer 407 combined with the third formulation, (5) Freeze dried Liposomal DOTAP dissolved in Sorensen's Buffer Solution, (6) Freeze dried Liposomal DOTAP combined with poloxamer 407 dispersed in Sorensen's Buffer Solution, (7) Freeze dried Liposomal DOTAP combined with the third formulation, and (8) Lysozyme and Lactoferrin dissolved in Sorensen's Buffer Solution, which was then incorporated into poloxamer 407 and combined with Freeze dried Liposomal DOTAP. The positive and negative control groups were 0.2% chlorhexidine gel and empty hydroxyapatite discs, respectively. Statistical evaluation was carried out with Kruskal-Wallis and Dunn's multiple comparison tests., Results: It was observed that the first, third and fifth groups did not have any antibacterial effects on the tested bacteria. The groups that contained poloxamer 407 had nearly identical antibacterial effects on Mutans Streptococci and L. acidophilus. These formulations also inhibited biofilm formation of the bacteria (p<0.05) more effectively. In the positive control group, there was no biofilm formation., Conclusions: Among the formulations containing poloxamer 407, the one containing Lysozyme and Lactoferrin exhibited the highest inhibitory effect on the tested bacteria. This novel formulation can be beneficial as an antibacterial agent for the prevention of dental caries and biofilm formation., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Streptococcus mutans and Streptococcus sobrinus are the major causative agents of human dental caries. Therefore, the removal or inhibition of these streptococcal biofilms is essential for dental caries prevention. In the present study, we evaluated the effects of ribose treatment alone or in combination with xylitol on streptococcal biofilm formation for both species. Furthermore, we examined the expression of genes responsible for dextran-dependent aggregation (DDAG). In addition, we investigated whether ribose affects the biofilm formation of xylitol-insensitive streptococci, which results from long-term exposure to xylitol. The viability of streptococci biofilms formed in a 24-well polystyrene plate was quantified by fluorescent staining with the LIVE/DEAD bacterial viability and counting kit, which was followed by fluorescence activated cell sorting analysis. The effects of ribose and/or xylitol on the mRNA expression of DDAG-responsible genes, gbpC and dblB, was evaluated by RT-qPCR. Our data showed that ribose and other pentose molecules significantly inhibited streptococcal biofilm formation and the expression of DDAG-responsible genes. In addition, co-treatment with ribose and xylitol decreased streptococcal biofilm formation to a further extent than ribose or xylitol treatment alone in both streptococcal species. Furthermore, ribose attenuated the increase of xylitol-insensitive streptococcal biofilm, which results in the reduced difference of biofilm formation between S. mutans that are sensitive and insensitive to xylitol. These data suggest that pentose may be used as an additive for teeth-protective materials or in sweets. Furthermore, ribose co-treatment with xylitol might help to increase the anti-cariogenic efficacy of xylitol., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Objectives: The antibacterial activity of Coffea canephora extract was evaluated in vitro against Streptococcus mutans and Streptococcus sobrinus. The viability of planktonic cells was analysed by susceptibility tests (MIC and MBC) and time-kill assays. The effect of the extract on dental demineralisation was also investigated., Methods: Primary 1st molar fragments (n=24) were inoculated with a saliva pool and sustained in a multiple plaque growth system for 10 days to form biofilm. The biofilm was treated with light roasted C. canephora extract at 20%, Milli-Q water (negative control) and chlorhexidine (positive control) once a day, during a week. Blank controls comprised fragments without treatment. Biofilm pH was monitored in the last day of treatment. Changes in tooth mineralisation were assessed by cross-sectional microhardness (CSMH) test., Results: MIC and MBC for S. mutans were 7±2 mg/mL and 160±0 mg/mL, respectively, showing no activity for S. sobrinus. The extract produced a 4-log reduction in the number of colonies of S. mutans after 3-h treatment (p<0.05) with undiluted extract (20%) and MBC concentration (16%). There was no difference among negative/blank controls and coffee plaque pH. Differences between CSMH values of dental fragments subjected to the coffee extract and to chlorhexidine were not significant. At depths up to 30 μm from the enamel surface, coffee extract and chlorhexidine promoted higher CSMH values when compared to blank/negative controls (p<0.05)., Conclusion: Our data suggest that light roasted C. canephora extract is beneficial as an anticariogenic substance., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Objectives: To determine the minimal inhibitory concentrations (MICs) of bacteriocin PsVP-10, chlorhexidine and triclosan on S. mutans and S. sobrinus and to study the potential synergistic combination between these antimicrobial and the bacteriocin PsVP-10., Design: Were determined MICs of bacteriocin PsVP-10, triclosan and chlorhexidine on strains of S. mutans and S. sobrinus, which formed a biofilm or did not form a biofilm. In addition, the synergistic effect was analysed by the determination of respective fractionary inhibitory concentrations (FICs) between bacteriocin PsVP-10 plus chlorhexidine and bacteriocin PsVP-10 plus triclosan., Results: MICs of three antimicrobials used were higher in those bacterial strains, which formed a biofilm. An interesting synergistic effect on both studied species was observed when bacteriocin and chlorhexidine were combined. A slighter synergy was determined for the combination bacteriocin PsVP-10 and triclosan., Conclusions: The results showed that the combination of chlorhexidine bacteriocin PsVP-10 could reduce the number of cariogenic bacteria for in vitro studies. In the future this synergistic combination could be an alternative to antimicrobial therapy against S. mutans or S. sobrinus.

Objectives: Polygonum cuspidatum has been used in Korean folk medicine to improve oral hygiene. This study was performed to evaluate the effects of methanol extract from root of P. cuspidatum (MEP) on bacterial viability and the virulence factors of Streptococcus mutans and Streptococcus sobrinus., Methods: To test the effects of MEP on bacterial viability, we determined the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against 20 bacterial strains, including S. mutans and S. sobrinus, using a micro-dilution assay. In case of S. mutans and S. sobrinus, the assays for time-kill and bacterial growth rate at sub-MIC concentrations were also performed. To determine effects of the extract on the virulence factors of S. mutans and S. sobrinus, the assays for sucrose-dependent adherence, water-insoluble glucan formation, glycolytic acid production, and acid tolerance were performed at sub-MIC levels. Phytochemical analysis for constituents of MEP was carried out., Results: MEP showed a broad antibacterial range (MIC 0.5-4 mg/ml). The MBC was two to four times higher than the MIC. The time-kill curves showed S. mutans and S. sobrinus were significantly killed after 1h of incubation. At sub-MIC levels, doubling times of S. mutans and S. sobrinus dose-dependently increased up to 211% and 123%, respectively. At sub-MIC levels, MEP also showed inhibitory effects on the virulence factors of S. mutans and S. sobrinus in a dose-dependent fashion. Phytochemical analysis revealed the presence of alkaloids, sterol/terpenes, tannins, flavonoids, and carbohydrates., Conclusion: These data indicate that MEP has inhibitory effects on bacterial viability at higher concentrations (> or =MIC) and the virulence factors of S. mutans and S. sobrinus at sub-MIC concentrations, suggesting that it might be useful for the control of dental plaque formation and subsequent dental caries formation.

Chocolate is suspected to contain some caries-inhibitory substances. The cariostatic activity of cacao mass extract (CM), the main component of chocolate, was examined in vitro and in experimental animals. CM showed no detectable effects on the cellular growth and acid production of mutans streptococci. On the other hand, the cell-surface hydrophobicity of mutans streptococci was significantly reduced by the presence of CM. Furthermore, insoluble glucan synthesis by the glucosyltransferases from either Streptococcus mutans MT8148R or Strep. sobrinus 6715 was inhibited by CM, but not significantly. Hence, the sucrose-dependent cell adherence of mutans streptococci was also depressed by CM. Finally, CM in both a 40% sucrose diet and drinking water resulted in reductions of caries development and plaque accumulation in rats infected with Strep. sobrinus 6715, but not significantly. These results indicate that cacao mass extract possesses some anticariogenic potential, but its anticaries activity is not strong enough to suppress significantly the cariogenic activity of sucrose.

Cacao bean husk extract (CBH) was examined for inhibitory effects on the caries-inducing properties of mutans streptococci in vitro and on caries development in specific pathogen-free Sprague-Dawley rats infected with mutans streptococci. CBH reduced the growth rate of almost all oral streptococci examined, which resulted in the reduction of acid production. Furthermore, insoluble glucan synthesis by the glucosyltransferases from Streptococcus mutans MT8148R and Streptococcus sobrinus 6715 was significantly inhibited by CBH. Hence, the sucrose-dependent cell adherence of mutans streptococci was also depressed by CBH. The administration of CBH in drinking water resulted in significant reductions of caries development and dental plaque accumulation in rats infected with either Strep. sobrinus 6715 or Strep. mutans MT8148R, and the minimum cariostatic concentration was 1.0 mg/ml. These results indicate that CBH possesses powerful anticariogenic potential.

Arnica and propolis have been used for thousands of years in folk medicine for several purposes. They possess several biological activities such as anti-inflammatory, antifungal, antiviral and tissue regenerative, among others. Although the antibacterial activity of propolis has already been demonstrated, very few studies have been done on bacteria of clinical relevance in dentistry. Also, the antimicrobial activity of Arnica has not been extensively investigated. Therefore the aim here was to evaluate in vitro the antimicrobial activity, inhibition of adherence of mutans streptococci and inhibition of formation of water-insoluble glucan by Arnica and propolis extracts. Arnica montana (10%, w/v) and propolis (10%, w/v) extracts from Minas Gerais State were compared with controls. Fifteen microorganisms were used as follows: Candida albicans--NTCC 3736, F72; Staphylococcus aureus--ATCC 25923; Enterococcus faecalis--ATCC 29212; Streptococcus sobrinus 6715; Strep. sanguis--ATCC 10556; Strep. cricetus--HS-6; Strep. mutans--Ingbritt 1600; Strep. mutans--OMZ 175; Actinomyces naeslundii--ATCC 12104, W 1053; Act. viscosus OMZ 105; Porphyromonas gingivalis; Porph. endodontalis and Prevotella denticola (the last three were clinical isolates). Antimicrobial activity was determined by the agar diffusion method and the zones of growth inhibition were measured. To assess cell adherence to a glass surface, the organisms were grown for 18 h at 37 degrees C in test-tubes at a 30 degree angle. To assay water-insoluble glucan formation, a mixture of crude glucosyltransferase and 0.125 M sucrose was incubated for 18 h at 37 degrees C in test-tubes at a 30 degree angle. Arnica and propolis extracts (20 microl) were added to these tubes to evaluate the % of inhibition of cell adherence and water-insoluble glucan formation. The propolis extract significantly inhibited all the microorganisms tested (p < 0.05), showing the largest inhibitory zone for Actinomyces spp. The Arnica extract did not demonstrate significant antimicrobial activity. Cell adherence and water-insoluble glucan formation were almost completely inhibited by the propolis extract at a final concentration of 400 microg/ml and 500 microg/ml, respectively. The Arnica extract showed slight inhibition of the adherence of the growing cells (19% for Strep. mutans and 15% for Strep. sobrinus) and of water-insoluble glucan formation (29%) at these same concentrations. Thus, the propolis extract showed in vitro antibacterial activity, inhibition of cell adherence and inhibition of water-insoluble glucan formation, while the Arnica extract was only slightly active in those three conditions.

Four groups of rats were inoculated with Streptococcus sobrinus ATCC 33478 and fed a cariogenic diet for 42 days. Topical treatment with either distilled water, sodium fluoride (0.2%), a solution containing lactoferrin and lactoperoxidase, or a solution containing liposome-encapsulated lactoferrin and liposome-encapsulated lactoperoxidase was applied at intervals for 35 days. Caries incidence in groups treated with liposome-encapsulated lactoferrin and lactoperoxidase was significantly lower than in control groups. The number of viable Strep. sobrinus and the proportion of Strep. sobrinus in the total counts were significantly higher in liposome-treated groups. Free lactoferrin and lactoperoxidase did not cause a significant reduction in caries incidence.

Cell suspensions of Streptococcus sobrinus can be aggregated by high molecular-weight alpha-1,6 glucans. The aggregation depends on the fidelity of a cell wall-bound, glucan-binding lectin (GBL). It is thought that the lectin may play a part in the sucrose-dependent accretion of streptococci in dental plaques. Results showed that the anionic detergent, sodium dodecyl sulphate (SDS) was a potent inhibitor of the lectin. When cells were incubated in SDS and washed to remove the detergent, lectin activity was diminished. Following incubation of the cells with SDS in the presence of glucan T-10, a low molecular-weight alpha-1,6 glucan, the loss of activity was less pronounced, suggesting that the glucan afforded partial protection against denaturation. Urea and guanidine hydrochloride were good inhibitors of the lectin, but, unlike SDS, were not able to inhibit it irreversibly, except at very high concentrations. Cationic detergents, such as cetylpyridinium bromide (and chloride), also irreversibly denatured the streptococcal lectin, but were not as effective as SDS in abolishing its activity. The results suggest that alpha-1,6 glucan stabilizes the GBL of S. sobrinus, rendering it more resistant to the effect of chaotropes. This may be one reason why dental plaques tend to resist detergents in dentrifices.

An antimicrobial agent, 3-(trimethoxysilyl)-propyldimethyloctadecyl ammonium chloride, was immobilized on silica. Interaction between the material (termed) OAIS) and various oral bacterial species were then studied. Seven species of Streptococcus and two Actinomyces were investigated for their ability to adhere to this biomaterial. Cell-surface hydrophobicity and zeta-potential were examined as well. Analysis of extracted hydrophobic proteins which adhered to OAIS revealed that the adherence of these micro-organisms was closely related to the hydrophobicity of their cell surfaces. The results of zeta-potential assays indicated that negative charge on the cell surface inhibited adherence to OAIS. Gel electrophoresis revealed that OAIS could absorb cell-surface hydrophobic proteins from all bacterial species tested. Preadsorption of hydrophobic components on the cell surface inhibited adherence of the Strep. mutans strain to OAIS in a dose-dependent manner. The results indicate that OAIS adsorption of these oral bacteria was dependent on the degree of hydrophobicity of their surfaces. A major component of this adherence was hydrophobic cell-surface proteins.

Previous studies have shown that fluoride (F) administered concomitantly with sucrose in drinking water, in the diet, or alone is an effective cariostatic agent. The purpose of the present study was to determine the minimum concentration of F co-crystallized with sugar that may be used to prevent dental caries in rats subjected to a severe cariogenic challenge. Desalivated Sprague-Dawley rats, infected with Streptococcus sobrinus, were placed in a König-Höfer programmed feeder. The rats received 17 meals daily for 21 days as follows: group (1) sucrose and sterile distilled water (s.d.w.); (2) sucrose and 1 part/10(6) F-water; (3) 1 part/10(6) F-sucrose and s.d.w.; (4) 4 parts/10(6) F-sucrose and s.d.w.; (5) 8 parts/10(6) F-sucrose and s.d.w.; (6) sucrose and 10 parts/10(6) F-water. Essential nutrition was administered by gavage. At death, blood was collected from each animal and one-half of the lower jaw was sonicated in 0.9% saline solution for microbial assessment and F analysis. Keyes smooth-surface and sulcal caries scores were significantly lower in the groups that received 10 parts/10(6) F-water and 8 parts/10(6) F-sucrose than in all other groups. The F concentration in the jaw suspension and plasma were significantly higher in the 10 parts/10(6) F-water and the 8 parts/10(6) F-sucrose groups than in all other groups. The total cultivable flora and Strep. sobrinus populations were lowest in F groups but this did not reach statistical significance. It is concluded that 8 parts/10(6) F co-crystallized with sucrose reduces the cariogenic potential of sugar as effectively as 10 parts/10(6) F in water, that is as little as 1 part/10(6) in sucrose has a significant effect, and that this cariostatic action is related to the amount of F in the oral environment.

The purpose was to explore the effects of combinations of copper, iron and fluoride (Cu, Fe and F) incorporated in sucrose by co-crystallization on caries development in the deslivated rat model and to examine acid formation by bacteria in the rat mouth. Ninety-six Sprague-Dawley rats were infected with Streptococcus sobrinus 6715 and desalivated when aged 26 days. Eight groups were placed in a König-Höfer programmed feeder and received 17 meals daily at hourly intervals, and essential nutrition (NCP No. 2) by gavage twice daily for 21 days. The groups received (1) plain sucrose, (2) F (8 parts/10(6)) co-crystallized with sucrose, (3) Fe (88 parts/10(6)) sucrose, (4) Cu (75 parts/10(6)) sucrose, (5) Cu + F sucrose, (6) Cu + L Fe sucrose, (7) F + Fe sucrose, and (8) Cu + Fe + F sucrose. At death the jaws were removed and sonicated in 0.9% saline solution for microbial assessment. In addition, organic acid assays were performed for each animal. Keyes smooth-surface and sulcal caries scores were lowest in the Cu + Fe + F sucrose group, but not statistically significantly different from those of the other Cu groups. The numbers of Strep. sobrinus found in the groups that received Cu, Cu + Fe, Cu + F, F + Fe and Cu + Fe + F sugar were lower than in the control group. Lactic acid was found in lower concentrations in Fe, Cu, Cu + F, Cu + Fe and F + Fe groups than in the other groups. It appears that combinations of Cu; Fe and F co-crystallized with sugar may have an additive effect in reducing the cariogenic potential of sugar by affecting lactic acid formation and reducing bacterial colonization.

Two experiments were conducted to find out what concentration of fluoride (F), when added as NaF to the fluid of an in vitro caries model, would be sufficient for full protection of the underlying enamel during a long (20 or 40 h) fall in 'plaque' pH. An existing caries model with bovine enamel and Streptococcus sobrinus 'plaque' was employed, while the fluid phase was initially either fully or partially saturated with tricalcium phosphate. In the model with fully saturated and more strongly buffered fluid, a 10 parts/10(6) addition of F to the fluid phase caused no inhibition of the pH fall. In the model with partially saturated and weakly buffered fluid, prevention of the pH fall increased along with increase of added F from 2 to 20 parts/10(6). In each model, the 10 parts/10(6) addition of F was associated with complete prevention of the increase of calcium (Ca) in the fluid phase. In the partially saturated, weakly buffered model, findings on the inorganic P of the fluid phase agreed with findings on Ca. The findings for microhardness of the enamel surface indicated a complete prevention of the enamel softening through the 10 parts/10(6) F additive. Enamel F content was elevated by the fermentation, and even more so when F was added with sugar to the fluid phase. The Ca content of Strep. sobrinus cells was reduced when incubated with sucrose for 40 h and, especially, with sucrose and F.(ABSTRACT TRUNCATED AT 250 WORDS)

The ability of various forms of human lactoferrin (LF) to agglutinate oral Streptococcus mutans, Strep. sobrinus, Strep. rattus, Strep. sanguis, Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans cells was studied spectrophotometrically. Fe3+ saturated LF was unable to agglutinate these bacteria, whereas iron-free LF (apo LF) effectively agglutinated Strep. mutans cells but not the other bacteria. The efficiency and rate of agglutination of Strep. mutans were somewhat lower with apo LF than with human whole saliva. However, secretory IgA, phosphate and whole saliva almost totally abolished the apo LF-mediated agglutination of Strep. mutans, suggesting binding to the same target sites on bacterial cell surfaces, or to each other. The presence of exogenous iron (Fe2+, Fe3+), lactoperoxidase or serum albumin did not affect the agglutination by apo LF. Low Ca2+ (50-100 microns) slightly enhanced the agglutination by apo LF but higher concentrations (0.5-1.0 mM) totally blocked the apo LF-mediated agglutination of Strep. mutans. Both saliva and apo LF significantly delayed the rapid autoaggregation of P. gingivalis cells. Aggregation of P. gingivalis is considered a potential virulence factor and a protective mechanism against the host's cellular defences in the gingival crevice. These findings show a novel, strain-specific antibacterial mechanism for LF against Strep. mutans and P. gingivalis and adds a new compound to the group of agglutinating proteins in human saliva.